Protein kinases C translocation responses to low concentrations of arachidonic acid

Arachidonic acid (AA) directly activates protein kinases C (PKC) and may th ereby serve as a regulatory signal during cell stimulation. The effect, how ever, requires a greater than or equal to 20 muM concentration of the fatty acid. We find that human polymorphonuclear neutrophils (PMN) equilibrated with a ligand for the diacylglycerol receptor on PKC, [H-3]phorbol dibutyra te (PDB), increased binding of [3H]PDB within 15 s of exposure to greater t han or equal to 10-30 nM Ak Other unsaturated fatty acids, but not a satura ted fatty acid, likewise stimulated PDB binding. These responses, similar t o those caused by chemotactic factors, resulted from a rise in the number o f diacylglycerol receptors that were plasma membrane-associated and therefo re accessible to PDB. Unlike chemotactic factors, however, AA was fully act ive on cells overloaded with Ca2+ chelators, The major metabolites of AA ma de by PMN, leukotriene B-4 and 5-hydroxyicosatetraenoate, did not mimic Ak and an AA antimetabolite did not block responses to Ak AA also induced PMN to translocate cytosolic PKC alpha, beta (II), and delta to membranes. This response paralleled PDB binding with respect to dose requirements, time, C a2+-independence, resistance to an AA antimetabolite, and induction by anot her unsaturated fatty acid but not by a saturated fatty acid. Finally, HEK 293 cells transfected with vectors encoding PKC beta (1) or PKC delta fused to the reporter enhanced green fluorescent protein (EGFP) were studied, AA caused EGFP-PKC beta translocation from cytosol to plasma membrane at grea ter than or equal to0.5 muM, and EGFP-PKC delta translocation from cytosol to nuclear and, to a lesser extent, plasma membrane at as little as 30 nM. We conclude that AA induces PKC translocations to specific membrane targets at concentrations 2-4 orders of magnitude below those activating the enzym es. These responses, at least as they occur in PMN, do not require changes in cell Ca2+ or oxygenation of the fatty acid. AA seems more suited for sig naling the movement than activation of PKC.